ABSTRACT While Alzheimer's disease (AD) has a high heritability, mutations in a few genes in early-onset familial AD account for only ~5% of cases. By comparison, the genetic architecture of late-onset AD (LOAD), which represents the majority of cases, is much more complex. Recent genome-wide association studies (GWAS) new provide insight into this complexity with the identification of more than 20 new genes that contribute to a large fraction of LOAD risk. However, a critical knowledge gap and a major challenge to translating these discoveries into treatments is that the neurobiology of most of these LOAD risk genes is not well understood. ABCA7, role these Remarkably, several LOAD genes, including BIN1, PICALM, APOe4, EPHA1, CD2AP and SORL1, have been previously impl icated in endosomal trafficking supporting an important for endosomal trafficking in LOAD. In addition, exome sequencing studies have identified rare coding mutations in genes enriched in patients with LOAD, which could provide insight into their function.While recent evidence implicates glutamatergic synapses as key pathogenic sites in AD, particularly in early stages of disease progression, the roles of LOAD risk genes in postsynaptic trafficking have not yet been investigated. Among these, the BIN1 gene was identified in several replicated GWAs as the most important risk locus for LOAD after APOE. Our preliminary bioinformatics analysis supports a prominent role for postsynaptic trafficking in LOAD, with a central role for Bin1. Our studies further show that for the first time that Bin1 is integral to the maintenance of postsynaptic membrane traffic, dendritic spine morphology, and AMPA receptor-mediated synaptic transmission. Bin1 is also known to interact with BACE1 and regulate APP processing. Using super-resolution and immuno-electron microscopy, we found that Bin1 is abundant at postsynaptic sites, where it is associated with several discrete compartments, including trafficking vesicles, the postsynaptic density, and cytoskeleton. We found that Bin1 forms protein complexes with the trafficking protein Arf6 and glutamate receptor subunit GluA1 in the rat brain, and modulates their trafficking in spines. In addition, we detected several other LOAD risk gene-encoded proteins at synapses. Because several LOAD risk genes have putative roles at synapses and in endosomal trafficking, we propose to define the functions of LOAD risk genes in postsynaptic endosomal trafficking at glutamatergic synapses. (Vassar), Here we and neuroprtoteomics (Savas), and will combine our expertise in synapse biology (Penzes), AD biology will employ advanced imaging, including super-resolution and in vivo two- photon microscopy, in combination with molecular, neuroproteomics, and electrophysiological, approaches, to test the hypothesis that a subset of LOAD risk genes modulate endosomal trafficking to regulate glutamate receptor and APP trafficking at postsynaptic sites. Their rare coding mutations may contribute to synapse pathogenesis. We will test this hypothesis in the following Aims: 1) To determine the mechanisms of regulation of postsynaptic trafficking by Bin1; 2) To define the mechanisms of regulation of dendritic and spine architecture by Bin1; 3) Regulation of spiny synapse function and architecture by Bin1 in the intact brain. The overarching goal of this proposal is to understandthe role ofsynaptic sites in trafficking in LOAD by determining the trafficking functions of major LOAD risk genes at synapses, critical AD pathogenesis. The proposed studies are highly innovative and impactful, because they will investigate the functions in synaptic trafficking of LOAD risk factors using state-of-the-art imaging tools be the first to such as super- resolution and two-photon imaging, and will uncover their functional interactions at synapses. Because the proposed work is at the convergence of several highly dynamic fields?synaptic biology, advanced imaging, neuroproteomics, and neurogenetics?it has a strong potentialto create new scientific paradigms.